Effect of the hepatitis C virus non-structural protein 4B on the levels of YAP transcription, expression, phosphorylation and intracellular distribution

ObjectiveTo construct the intracellular fluorescent expression plasmid of non-structural protein 4B(NS4B) of hepatitis C virus(HCV), and detect the effects of NS4B on the levels of transcription, expression, phosphorylation and intracellular distribution of Yes-associated protein(YAP).

MethodsThe specific amplification primers for NS4B were designed according to the existing plasmid containing HCV 1b virus NS4B complete gene.The complete sequence of NS4B was amplified to construct the NS4B fluorescent plasmid, and the plasmid was transfected into HEPG2 and 293T cells.The transcription level of YAP gene, and expression and phosphorylation levels of YAP protein were detected using Real-time PCR and Western blotting, respectively.The intracellular distribution of YAP was detected by confocal laser and nucleo-cytoplasmic separation assay.

ResultsThe complete NS4B sequence was successfully amplified, and its fluorescent plasmid was constructed.The results of Real-time PCR and Western blotting showed that the NS4B did not affect the transcription and expression levels of YAP, but could regulate its phosphorylation level.The results of laser confocal and nucleo-cytoplasmic separation results suggested that the NS4B could regulate the ratio of YAP in cytoplasm and nucleus.

ConclusionsThe HCV NS4B may play a biological role through its effects on YAP.